Usnic acid

Since its first isolation in 1844, usnic acid [2,6-diacetyl-7,9-dihydroxy-8,9b-dimethyl-1,3(2H,9bH)-dibenzo-furandione] has become the most extensively studied lichen metabolite and one of the few that is commercially available. Usnic acid is uniquely found in lichens, and is especially abundant in genera such as Alectoria, Cladonia, Usnea, Lecanora, Ramalina and Evernia. Many lichens and extracts containing usnic acid have been utilized for medicinal, perfumery, cosmetic as well as ecological applications. Usnic acid as a pure substance has been formulated in creams, toothpaste, mouthwash, deodorants and sunscreen products, in some cases as an active principle, in others as a preservative. In addition to antimicrobial activity against human and plant pathogens, usnic acid has been shown to exhibit antiviral, antiprotozoal, antiproliferative, anti-inflammatory and analgesic activity. Ecological effects, such as antigrowth, antiherbivore and anti-insect properties, have also been demonstrated. A difference in biological activity has in some cases been observed between the two enantiomeric forms of usnic acid. Recently health food supplements containing usnic acid have been promoted for use in weight reduction, with little scientific support. The emphasis of the current review is on the chemistry and biological activity of usnic acid and its derivatives in addition to rational and ecologically acceptable methods for provision of this natural compound on a large scale.     

The comparison of miRNAs that respond to anti-breast cancer drugs and usnic acid for the treatment of breast cancer

This study was designed to compare usnic acid with anti-breast cancer drug molecules (A-BCDM) routinely used in the treatment of breast cancer. The miRNA information of 17 anti-breast cancer drug used in breast cancer treatment was obtained from the Small Molecule-miRNA Network-Based Inferance (SMIR-NBI) tool. We had been determined common and different expressed miRNAs between 17 A-BCDM & usnic acid and were classified according to the common miRNAs to reveal molecular similarity. As a result of the bioinformatic analyzes, 20 common miRNAs were determined between 17 A-BCDM and usnic acid. The common miRNAs were analyzed with bioinformatic tolls for determining pathways and targets. The most common miRNAs for 6 of 17 A-BCDM and usnic acid were determined as miR-374a-5p and miR-26a-5p. We compared the anti-proliferative effect of usnic acid and one of the 17 A-BCDM that tamoxifen on MDA-MB-231 triple negative breast cancer cell with real-time cell analysis system. The real time PCR assay was carried out with miR-26a-5p for evaluate to expression level of MDA-MB-231 breast cancer cell and MCF-12A non-cancerous epithelial breast cell. As a result of study, usnic acid as novel candidate drug molecule showed high similarity ratio with 5-Fluorouracil, Sulindac Sulfide, Curcumin and Cisplatin A-BCDM used in treatment of breast cancer. miR-26a-5p as common response miRNA of usnic acid and tamoxifen was showed a decreased level of expression by validated qRT-PCR assay. The obtained from study, in addition to 17 A-BCDM, usnic acid has also the potential to be used as a candidate molecule in the treatment of breast cancer. Moreover, miR-26a-5p might be used as a biomarker in the treatment of breast cancer but further analysis is required.     

Spatial trends in usnic acid concentrations of the lichen<Emphasis Type="Italic"> Flavocetraria nivalis</Emphasis> along local climatic gradients in the Arctic (Kongsfjorden,Svalbard)

The dibenzofuran usnic acid is an important secondary lichen metabolite, having many postulated biological roles, but evidence from field surveys is scanty. Trends in usnic acid concentrations and lobe width in the arctic lichen Flavocetraria nivalis were analysed along local longitudinal and altitudinal gradients in the Kongsfjorden area, northwestern Spitsbergen. Temperature was measured along the same gradients during 1 year. Other environmental variables were also estimated. The model that best explains the variability in usnic-acid levels includes the parameters effective temperature sum, frost sum and temperature range. Temperature range indicates a relationship between high usnic acid levels and humidity, whereas the two first parameters indicate a relationship with low temperatures, which could be direct or indirect. Much of the variability in usnic acid levels and lobe width could not be explained by the selected models. Thus, the secondary metabolism and lobe growth in this lichen is a complex matter, involving numerous environmental and possibly also intrinsic factors.     

Anti-viral activity of (−)- and (+)-usnic acids and their derivatives against influenza virus A(H1N1)2009

Influenza is a widespread respiratory infection. Every year it causes epidemics, quickly spreading from country to country, or even pandemics, involving a significant part of the human population of the earth. Being a highly variable infection, influenza easy accumulates the resistance mutations to many antivirals.Usnic acid, a dibenzofuran originally isolated from lichens belongs to the secondary metabolites and has a broad spectrum of biological activity. In humans, it can act as an anti-inflammatory, antimitotic, antineoplasic, antibacterial, and antimycotic agent. In this work we studied for the first time the antiviral activity of usnic acid and its derivatives against the pandemic influenza virus A(H1N1)pdm09. A total of 26 compounds representing (+) and (?) isomers of usnic acid and their derivates were tested for cytotoxicity and anti-viral activity in MDCK cells by microtetrazolium test and virus yield assay, respectively. Based on the results obtained, 50% cytotoxic dose (CTD₅₀) and 50% effective dose (ED₅₀) and selectivity index (SI) were calculated for each compound. Eleven of them were found to have SI higher than 10 (highest value 37.3). Absolute configuration was shown to have critical significance for the anti-viral activity. With minor exceptions, in the pair of enantiomers, (?)-usnic acid was more active comparing to (+)-isomer, but its biological activity was reversed after the usnic acid was chemically modified. Based on the obtained results, derivatives of usnic acid should be considered as prospective compounds for further optimization as anti-influenza substances.     

Usnic Acid Potassium Salt: An Alternative for the Control of Biomphalaria glabrata (Say, 1818)

In Brazil, the snail Biomphalaria glabrata is the most important vector of schistosomiasis due to its wide geographical distribution, high infection rate and efficient disease transmission. Among the methods of schistosomiasis control, the World Health Organization recommends the use of synthetic molluscicides, such as niclosamide. However, different substances of natural origin have been tested as alternatives for the control or eradication of mollusks. The literature describes the antitumor, antimicrobial and antiviral properties of usnic acid as well as other important activities of common interest between medicine and the environment. However, usnic acid has a low degree of water solubility, which can be a limiting factor for its use, especially in aquatic environments, since the organic solvents commonly used to solubilize this substance can have toxic effects on aquatic biota. Thus, the aim of the present study was to test the potassium salt of usnic acid (potassium usnate) with regard to molluscicidal activity and toxicity to brine shrimp (Artemia salina). To obtain potassium usnate, usnic acid was extracted with diethyl ether isolated and purified from the lichen Cladonia substellata. Biological assays were performed with embryos and adult snails of B. glabrata exposed for 24 h to the usnate solution solubilized in dechlorinated water at 2.5; 5 and 10 µg/ml for embryos, 0.5; 0.9; 1;5 and 10 µg/ml for mollusks and 0.5; 1; 5; 10 µg/ml for A. salina. The lowest lethal concentration for the embryos and adult snails was 10 and 1 µg/ml, respectively. No toxicity to A. salina was found. The results show that modified usnic acid has increased solubility (100%) without losing its biological activity and may be a viable alternative for the control of B. glabrata.     

Effect of (+)-usnic acid on mitochondrial functions as measured by mitochondria-specific oligonucleotide microarray in liver of B6C3F1 mice

Usnic acid is a lichen metabolite used as a weight-loss dietary supplement due to its uncoupling action on mitochondria. However, its use has been associated with severe liver disorders in some individuals. Animal studies conducted thus far evaluated the effects of usnic acid on mitochondria primarily by measuring the rate of oxygen consumption and/or ATP generation. To obtain further insight into usnic acid-mediated effects on mitochondria, we examined the expression levels of 542 genes associated with mitochondrial structure and functions in liver of B6C3F₁ female mice using a mitochondria-specific microarray. Beginning at 8 weeks of age, mice received usnic acid at 0, 60, 180, and 600 ppm in ground, irradiated 5LG6 diet for 14 days. Microarray analysis showed a significant effect of usnic acid on the expression of several genes only at the highest dose of 600 ppm. A prominent finding of the study was a significant induction of genes associated with complexes I through IV of the electron transport chain. Moreover, several genes involved in fatty acid oxidation, the Krebs cycle, apoptosis, and membrane transporters were over-expressed. Usnic acid is a lipophilic weak acid that can diffuse through mitochondrial membranes and cause a proton leak (uncoupling). The up-regulation of complexes I–IV may be a compensatory mechanism to maintain the proton gradient across the mitochondrial inner membrane. In addition, induction of fatty acid oxidation and the Krebs cycle may be an adaptive response to uncoupling of mitochondria.     

Target network analysis of adiponectin,a multifaceted adipokine

Application of network analysis to dissect the potential molecular mechanisms of biological processes and complicated diseases has been the new trend in biology and medicine in recent years. Among which, the protein–protein interactions (PPI) networks attract interests of most researchers. Adiponectin, a cytokine secreted from adipose tissue, participates in a number of metabolic processes, including glucose regulation and fatty acid metabolism and involves in a series of complicated diseases from head to toe. Hundreds of proteins including many identified and potential drug targets have been reported to be involved in adiponectin related signaling pathways, which comprised a complicated regulation network. Therapeutic target database (TTD) provides extensive information about the known and explored therapeutic protein targets and the signaling pathway information. In this study, adiponectin associated drug targets based PPI was constructed and its topological properties were analyzed, which might provide some insight into the dissection of adiponectin action mechanisms and promote adiponectin signaling based drug target identification and drug discovery. J. Cell. Biochem. 114: 1145–1152, 2013. © 2012 Wiley Periodicals, Inc.     

Novel derivatives of usnic acid effectively inhibiting reproduction of influenza A virus

Influenza virus is serious human pathogen leading to high morbidity and mortality all over the world. Due to high rate of mutation, it is able to fast development of drug resistance that makes necessary to search novel antivirals with broad range and alternative targets. In the present study we describe synthesis and anti-viral activity of novel derivatives of usnic acid (2,6-diacetyl-7,9-dihydroxy-8,9b-dimethyl-1,3(2H,9bH)-dibenzo-furandione). It is shown that anti-viral activity of usnic acid can be increased by side moieties introduction. The modification with chalcones appeared to be the most effective. Our study revealed that (−)-usnic acid exhibited higher antiviral activity than its (+)-enantiomer, but in the pairs of enantiomer derivatives such as enamines, pyrazoles and chalcones, the (+)-enantiomers were more potent inhibitors of the virus. For other groups of compounds the inhibiting activities of the enantiomers were comparable. Further optimization of the structure could therefore result in development of novel anti-influenza compound with alternative target and mechanism of virus-inhibiting action.     

Evaluation of Anticancer and Antioxidant Activity of a Commercially Available CO2 Supercritical Extract of Old Man’s Beard (Usnea barbata)

There is a worldwide ongoing investigation for novel natural constituents with cytotoxic and antioxidant properties. The aim of this study was to investigate chemical profile and stated biological activities of the supercritical CO₂ extract (SCE) of old man’s beard compared to the extracts obtained using the conventional techniques (Soxhlet extracts and macerate). The most abundant compound identified was usnic acid, which content was inversely proportional to the polarity of the solvent used and was the highest in the SCE, which was the sample revealing the highest cytotoxic activity in tested tumor cell lines (B16 mouse melanoma and C6 rat glioma), with lower IC₅₀ values compared to pure usnic acid. Further investigations suggested both SCE and usnic acid to induce apoptosis and/or autophagy in B16 and C6, indicating higher cytotoxicity of SCE to be related to the higher degree of ROS production. A good correlation of usnic acid content in the extracts and their antioxidant capacity was established, extricating SCE as the most active one. Presented results support further investigations of SCE of old man’s beard as a prospective therapeutic agent with potential relevance in the treatment of cancer and/or in oxidative stress-mediated conditions.     

DNA水凝胶的制备及应用

脱氧核糖核酸(Deoxyribonucleic acid,DNA)不仅可作为生物遗传的物质基础,又以其可编程性、功能多样性、生物相容性和生物可降解性等优点,在生物材料的构建方面表现出巨大的潜力。DNA水凝胶是一种主要由DNA参与形成的三维网状聚合物材料,同时因其保留的DNA生物性能与自身骨架的机械性能的完美融合使得它成为近年来最受关注的新兴功能高分子材料之一。目前,基于各种功能核酸序列或通过结合不同的功能材料制备的单组分或多组分DNA水凝胶,已广泛用于生物医学、分子检测及环境保护的研究或应用领域中。文中主要总结了近十几年来DNA水凝胶制备方法上的研究进展,探讨了DNA水凝胶的分类策略,并进一步综述了DNA水凝胶在药物运输、生物传感、细胞培养等方面的应用研究。最后对DNA水凝胶未来的发展方向以及可能面临的挑战进行了展望。     

松萝酸提取工艺的研究

以破茎松萝为材料,分别以碱水或碱醇直接提取、制备松萝酸钠,并对松萝酸钠提取影响因素进行探究。对所得提取物进行分离和定性鉴定,并利用高效液相层析技术测定提取物中松萝酸的含量。通过单因素实验考察了NaOH和Na2CO3浓度对松萝酸提取的影响,用HPLC测定不同浓度碱液提取物中松萝酸的含量。实验结果说明,1.5%NaOH和75%乙醇+4%Na2CO3提取松萝酸效果较好,松萝酸提取得率最高,分别为1.75%和1.99%,破茎松萝材料中松萝酸含量为2.4%。     

Enantioselective activity of usnic acid: a comprehensive review and future perspectives

Phytochemistry Reviews - This review is focused on the comparison of the biological and pharmacological activities of usnic acid enantiomers. Most of the available data refer to (+)-usnic acid,...     

ANTIMICROBIAL,ANTIVIRAL AND CYTOTOXIC ACTIVITY OF NEW ZEALAND LICHENS

A total of 69 species of lichens have been collected from various locations around New Zealand. Screening of extracts of these species for antimicrobial, antiviral and cytotoxic activity showed a high proportion with biological activity. Active extracts were generally from species known to contain phenolic compounds. Bioactivity-directed isolation work onCladia retipora, Pseudocyphellaria glabra and P. homoeophylla led to the identification of usnic acid as the main antimicrobial, cytotoxic and antiviral component in these three species.     

Mycotoxins and usnic acid and their distribution in lichens belonging to the genera Cetraria,Flavocetraria, and Cladonia

The content and the pattern of distribution of mycotoxins and usnic acid in thalli of lichens belonging to the genera Cladonia (four species), Cetraria (two species), and Flavocetraria (two species) have been studied using the method of immunoenzymatic analysis. Statistically significant differences have been estimated between the upper and the lower parts of the thalli. The levels of the mycotoxins in the lower, older parts of the thalli have been higher than in the upper, younger ones. The comparison of lichen specimens belonging to the same species and collected in the same location at different years did not reveal statistically significant differences in the content of mycotoxins both in the upper and the lower parts of the thalli. Statistically significant differences in the usnic acid content have been revealed only in the case of C. stellaris. In the upper part of the podetia of this species, the usnic acid content has been higher than in the lower one.     

Proton-Shuttling Lichen Compound Usnic Acid Affects Mitochondrial and Lysosomal Function in Cancer Cells

The lichen compound usnic acid (UA) is a lipophilic weak acid that acts as a proton shuttle and causes loss of mitochondrial inner membrane potential. In the current study we show that UA treatment induced the formation of autophagosomes in human cancer cells, but had minimal effects on normal human fibroblasts. However, autophagic flux was incomplete, degradation of autophagosomal content did not occur and acidification was defective. UA-treated cells showed reduced ATP levels and activation of AMP kinase as well as signs of cellular stress. UA is thus likely to trigger autophagosome formation both by energy depletion and stress conditions. Our findings indicate that the H⁺-shuttling effect of UA operates not only in mitochondria as previously shown, but also in lysosomes, and have implications for therapeutic manipulation of autophagy and pH-determined drug distribution.     

Insulin 3: From chemical synthesis to biological function

Summary Insulin-like peptide 3 (INSL3) is one of ten members of the human insulin superfamily and consists of two peptide chains that contain the characteristic insulin fold and disulfide bond pairings. It is primarily produced in the Leydig cells of the testes, and gene knockout experiments have identified a key biological role as initiating testes descent during foetal development. Its receptor has recently been shown to be a member of the leucine-rich repeat-containing G-protein-coupled receptor family (LGR) and is known as LGR8. Considerable work has recently been undertaken with the aim of studying the mechanism of INSL3 downstream action on responsive cells and, towards this goal, the use of synthetic peptides has proved particularly beneficial. This mini-review outlines how these together with basic structure-function studies are beginning to reveal not only its molecular actions but also its potential new biological actions.     

4-Phenylbutyric acid promotes plant regeneration as an auxin by being converted to phenylacetic acid via an IBR3-independent pathway

4-Phenylbutyric acid (4PBA) is utilized as a drug to treat urea cycle disorders and is also being studied as a potential anticancer drug that acts via its histone deacetylase (HDAC) inhibitor activity. During a search to find small molecules that affect plant regeneration in Arabidopsis, we found that 4PBA treatment promotes this process by mimicking the effect of exogenous auxin. Specifically, plant tissue culture experiments revealed that a medium containing 4PBA enhances callus formation and subsequent shoot regeneration. Analyses with auxin-responsive or cytokinin-responsive marker lines demonstrated that 4PBA specifically enhances AUXIN RESPONSE FACTOR (ARF)-dependent auxin responses. Our western blot analyses showed that 4PBA treatment does not enhance histone acetylation in Arabidopsis, in contrast to butyric acid and trichostatin A, other chemicals often used as HDAC inhibitors, suggesting this mechanism of action does not explain the observed effect of 4PBA on regeneration. Finally, mass spectroscopic analysis and genetic approaches uncovered that 4PBA in Arabidopsis plants is converted to phenylacetic acid (PAA), a known natural auxin, in a manner independent of peroxisomal IBR3-related β-oxidation. This study demonstrates that 4PBA application promotes regeneration in explants via its auxin activity and has potential applications to not only plant tissue culture engineering but also research on the plant β-oxidation pathway.     

Mechanism of action of an old antibiotic revisited: Role of calcium ions in protonophoric activity of usnic acid

Usnic acid (UA), an old antibiotic and one of the first described mitochondrial uncouplers, has demonstrated many beneficial activities, such as antimicrobial, antiviral, antitumour and anti-inflammatory properties. Here, we performed a thorough investigation of effects of usnic acid and its analogues on artificial planar bilayer lipid membrane (BLM), rat liver mitochondria and bacteria. Surprisingly enough, all of the three hydroxyl groups of UA appeared to be involved in its proton-shuttling activity on BLM. We ascribed this fact to an ability of UA to form complexes with calcium ions, aiding it in cycling protons across the membrane. Actually, the addition of calcium ions markedly stimulated the UA-induced electrical current across BLM. By using the calcium ionophore A23187, we proved the involvement of calcium ions in the UA uncoupling action on isolated rat liver mitochondria. The calcium-chelating property of UA was demonstrated here by the method of extracting metal ions into a hydrophobic phase. Modification of any of the hydroxyl groups in UA dramatically reduced not only the UA-induced current across BLM and the UA-mediated calcium extraction, but also the uncoupling activity of UA in mitochondria and the inhibiting effect of UA on the growth of Bacillus subtilis. The ability of UA to cause dissipation of membrane potential in isolated liver mitochondria and bacterial cells was shown here for the first time. In view of the data obtained, the protonophoric activity of UA is considered to make a significant contribution to its antibacterial action.